1. Field of the Invention
The present invention relates to tissue culture media useful for the in vitro growth of cells.
2. Description of the Prior Art
It is well-known that animal and plant cells may be grown in vitro in liquid culture media, i.e., tissue culture; see e.g., Kruse et al, Academic Press, New York, N.Y., 1973, and Ham, R. G. and McKeehan, W. L., Methods in Enzymology, 58; 44-93 (1979). Such media usually contain a wide array of different components, including various nutrients and salts which promote the maximum growth of the cultured cells.
Cells grown in tissue culture are used for many different purposes; for example, for the production of enzymes, cell products, antibodies, or for the general testing of drugs, carcinogenic agents and the like. In vitro growth of animal cell lines has recently acquired new relevance with the development of cell fusion, and the preparation of hybridomas and their associated monoclonal antibodies.
The art has long established that one of the essential components for tissue culture media is bovine serum, most preferably fetal calf or newborn calf serum. These two types of serum lack high concentrations of components which inhibit cell growth, and contain undefined factors which support cell growth in vitro. The use of fetal calf serum, however, is troubled by a lack of sufficient supply, and poor characterization of its ingredients. Furthermore, costs for this type of serum have prevented the economic growth of cells containing such serum.
A number of fetal calf serum substitutes have been proposed. For example, Michl, U.S. Pat. No. 3,128,228, discloses a culture medium for the preparation of tissue cultures on the basis of serum protein fractions, and a nutrient solution containing nutrient salts, protein fission products, and particular amino acids, further sugars and vitamins or coenzymes. The serum substitute is derived from calf blood by coagulation, isolation of the serum, followed by a series of precipitation steps.
Bozicevich, U.S. Pat. No. 3,429,867 describes a so-called "Agamma" calf serum suitable for tissue cultures, prepared from calf serum by precipitation and acidification thereof.
Birch, U.S. Pat. No. 4,038,139 describes a culture medium containing swine serum and about 0.1% of a surfactant which inhibits the precipitation of protein. The swine serum of Birch is stated to support the growth of lymphoid cells giving superior yields to those obtained using fetal calf serum. Swine serum is also considerably less expensive and thus brings about a concomitant reduction in cost.
Gaeta, U.S. Pat. No. 3,122,476, describes a substitute fetal calf serum useful for the growth of normal human cells and other animal cells in vitro, prepared from the blood of immature calves by fractionation, isolation of the serum and separation therefrom of gamma-globulins and other toxic substances by ethyl alcohol precipitation.
The difficulties with one or more of these prior art sera is that extensive and unselective precipitation by salt, acid or organic solvents causes the removal of essential growth factors which render the resulting substitute sera effective for only relative short periods of time; i.e., some of these sera are unable to support cell growth over many generations. Furthermore, it is well-known that calf serum contains a number of toxins not present in fetal calf serum, which toxins tends to inhibit cell growth. An additional disadvantage encountered in some of these sera is the lack of complete standardization of components, which would provide controllable conditions for cell growth in tissue cultures.
A need, therefore, continues to exist for a standardized, well-characterized fetal calf serum-substitute derived from calf serum, which contains active growth ingredients and lacks cell growth inhibiting toxins.